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The aim of this study was to evaluate the effect of polymorphisms affecting the clopidogrel metabolism (CYP3A4 IVS10+12G/A and CYP2C19∗2) and the P2Y12 receptor (P2Y12 T744C) on modulating platelet function in acute coronary syndrome patients on dual antiplatelet treatment.Residual platelet reactivity (RPR) phenomenon on antiplatelet therapy requires clarification. P2Y12 T744C, CYP3A4 IVS10+12G/A and, in healthy individuals only, CYP2C19∗2 polymorphisms have been investigated; however, the influence on platelet reactivity in a large population of high-risk vascular patients on dual antiplatelet treatment has not yet been elucidated.A total of 1419 acute coronary syndrome patients on dual antiplatelet treatment were studied. Platelet function was evaluated by platelet-rich plasma aggregation. Electronic nanochips and restriction-fragment length polymorphism were used for analysis of polymorphisms.Only CYP2C19∗2, out of the three investigated polymorphisms, is associated with higher platelet reactivity. Carriers of the ∗2 allele had significantly higher platelet aggregation values after arachidonic acid (AA; P=0.043), 2 μmol/l adenosine 5′ diphosphate (ADP; P<0.0001) and 10 μmol/l ADP (P=0.001) stimuli. The genotype distribution of CYP2C19∗2 polymorphism significantly differed between patients with and without RPR, as evaluated by 10-μmol/l ADP-induced platelet aggregation (P=0.002) and by AA-induced platelet aggregation (P=0.045). At the multivariate linear regression analysis, the CYP2C19∗2 polymorphism remained a significant and independent risk factor for dual antiplatelet treatment variability.This study demonstrates, for the first time, that the ∗2 CYP2C19 allele is associated with higher platelet aggregability and RPR in high-risk vascular patients on dual antiplatelet treatment. These findings can have a significant impact on the future design of pharmacogenetic antiaggregant strategies for high-risk vascular patients on dual antiplatelet treatment. [ABSTRACT FROM AUTHOR]

Rationale: Abdominal aortic aneurysm (AAA) is a complex disease with both genetic and environmental risk factors. Together, 6 previously identified risk loci only explain a small proportion of the heritability of AAA., Objective: To identify additional AAA risk loci using data from all available genome-wide association studies., Methods and Results: Through a meta-analysis of 6 genome-wide association study data sets and a validation study totaling 10 204 cases and 107 766 controls, we identified 4 new AAA risk loci: 1q32.3 (SMYD2), 13q12.11 (LINC00540), 20q13.12 (near PCIF1/MMP9/ZNF335), and 21q22.2 (ERG). In various database searches, we observed no new associations between the lead AAA single nucleotide polymorphisms and coronary artery disease, blood pressure, lipids, or diabetes mellitus. Network analyses identified ERG, IL6R, and LDLR as modifiers of MMP9, with a direct interaction between ERG and MMP9., Conclusions: The 4 new risk loci for AAA seem to be specific for AAA compared with other cardiovascular diseases and related traits suggesting that traditional cardiovascular risk factor management may only have limited value in preventing the progression of aneurysmal disease., (© 2016 The Authors.)

Objective: Among plasma proteins, fibrinogen represents a major target of oxidative modifications. In patients with post-acute myocardial infarction (6 months after the acute event), fibrinogen oxidation-induced carbonyls and fibrinogen function were estimated using in vitro and ex vivo approaches. Fibrinogen structural features and clot architecture were also explored., Approach and Results: In 39 patients with post-acute myocardial infarction and 28 age-, sex-, and risk factor-matched controls, oxidative stress markers (in plasma and in purified fibrinogen fractions), thrombin-catalyzed fibrin polymerization, and plasmin-induced fibrin lysis were estimated. Circular dichroism spectra of purified fibrinogen extracts, electron microscopy, and differential interference contrast microscopy analyses of fibrin clots were also performed. Marked signs of oxidative stress in plasma (P<0.01 versus controls) and, correspondingly, an increased extent of fibrinogen carbonylation (3.5-fold over control values; P<0.01 versus controls) were observed in patients. Furthermore, fibrinogen fractions purified from patients exhibited significantly reduced clotting ability and decreased susceptibility to plasmin-induced lysis (P<0.01 versus controls). Alterations in fibrinogen secondary structure, as suggested by circular dichroism spectroscopy, and in fibrin clot architecture, as analyzed by electron and differential interference contrast microscopy, were also identified., Conclusions: Here, we report for the first time that patients with post-acute myocardial infarction present with an overall imbalance in redox status and marked fibrinogen carbonylation associated with altered fibrinogen function, thus suggesting a role for carbonylation as a direct mechanism of fibrinogen function. The observed features occur along with modifications in protein structure and in clot architecture., (© 2014 American Heart Association, Inc.)

Background and Purpose: Experimentally, matrix metalloproteinases (MMPs) play a detrimental role related to hemorrhagic transformation and severity of an ischemic brain lesion. Tissue-type plasminogen activator (tPA) enhances such effects. This study aimed to expand clinical evidence in this connection., Methods: We measured MMPs 1, 2, 3, 7, 8, 9, and tissue inhibitors of metalloproteinases 1, 2, 4 circulating level in blood taken before and 24 hours after tPA from 327 patients (mean age, 68.9±12.1 years; median National Institutes of Health Stroke Scale, 11) with acute ischemic stroke. Delta median values ([24 hours post tPA-pre tPA]/pre tPA) of each MMP or tissue inhibitors of metalloproteinase were analyzed across subgroups of patients undergoing symptomatic intracerebral hemorrhage, 3-month death, or 3-month modified Rankin Scale score 3 to 6., Results: Adjusting for major clinical determinants, only matrix metalloproteinase-9 variation proved independently associated with death (odds ratio [95% confidence interval], 1.58 [1.11-2.26]; P=0.045) or symptomatic intracerebral hemorrhage (odds ratio [95% confidence interval], 1.40 [1.02-1.92]; P=0.049). Both matrix metalloproteinase-9 and tissue inhibitors of metalloproteinase-4 changes were correlated with baseline, 24 hours, and 7 days National Institutes of Health Stroke Scale (Spearman P from <0.001 to 0.040)., Conclusions: Our clinical evidence corroborates the detrimental role of matrix metalloproteinase-9 during ischemic stroke treated with thrombolysis, and prompts clinical trials testing agents antagonizing its effects.

Purpose: To investigate atherosclerotic and thrombophilic risk factors in patients affected by acute ischemic and nonischemic central retinal vein occlusions (CRVOs)., Methods: One hundred and three patients with acute unilateral CRVO (41 ischemic and 62 nonischemic) were studied. The frequency of traditional cardiovascular risk factors was assessed, and the plasma levels of a variety of thrombophilic markers were measured. Univariate logistic regression was performed to determine risk factors for ischemic CRVO., Results: Arterial hypertension, hypercholesterolemia, postmethionine hyperhomocysteinemia (HHcy), elevated factor VIII, and reduced folic acid and B6 plasma levels were more frequent in patients with ischemic CRVO than in those with nonischemic CRVO (P = 0.030, P = 0.025, P = 0.011, P < 0.001, P < 0.001, and P = 0.044, respectively). Risk factors for ischemic CRVO were arterial hypertension (odds ratio [OR], 3.22; 95% confidence interval [CI], 1.13-9.21; P = 0.037), hypercholesterolemia (OR, 3.03; 95% CI, 1.06-8.65; P = 0.042), reduced folic acid levels (OR, 6.77; 95% CI, 1.59-28.79; P = 0.011), and elevated FVIII levels (OR, 6.17; 95% CI, 2.56-14.82; P < 0.001). Postmethionine HHcy was associated with low folic acid levels (r = -0.413; P = 0.007; OR, 9.33; 95% CI, 2.06-42.18; P = 0.005)., Conclusion: The results of the present study suggest that some atherosclerotic and thrombophilic risk factors may increase the risk of having an ischemic form of CRVO.

Background and Purpose: Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is an inherited disease due to cerebral microangiopathy presenting with variable pictures, including stroke, progressive cognitive impairment, and disability. Mechanisms leading from vessel structural changes to parenchymal damage and eventually to clinical expression are not fully understood. Among pathogenic processes, endothelial dysfunction has been hypothesized. Endothelial progenitor cells and circulating progenitor cells (CPCs) derived from bone marrow participate in endothelium structure and function maintenance and contribute to ischemic area revascularization. No data are available about these cells in CADASIL. Our objective in this study was to evaluate endothelial progenitor cells and CPCs role in CADASIL., Methods: Twenty-nine patients with CADASIL and 29 sex- and age-matched control subjects were enrolled. Cells were measured in peripheral blood using flow cytometry. Endothelial progenitor cells were defined as positive for CD34/KDR, CD133/KDR, and CD34/CD133/KDR; and CPCs as positive for CD34, CD133, and CD34/CD133., Results: Endothelial progenitor cells were significantly lower in patients with CADASIL than in control subjects (CD34/KDR: 0.05 versus 0.1 cells/microL, P=0.005; CD133/KDR: 0.07 versus 0.1 cells/microL, P=0.006; CD34/CD133/KDR: 0.05 versus 0.1 cells/microL, P=0.001). The difference remained significant after adjusting for age, sex, and statin use. CPCs were not significantly lower in CADASIL, but patients with stroke or dementia had significantly reduced CPC levels than patients without (CD34: 1.68 versus 2.95 cells/microL, P=0.007; CD133: 1.40 versus 2.82 cells/microL, P=0.004; CD34/CD133: 1.44 versus 2.75 cells/microL, P=0.004). CPC levels significantly correlated with cognitive and motor performance measures., Conclusions: We have documented an association between endothelial progenitor cells and CPCs and CADASIL, extending previous data about the presence of endothelial dysfunction in this disease and its potential role in modulating phenotype.

Background: No data exist about the impact of high residual platelet reactivity (HRPR) after clopidogrel loading on long-term clinical outcome in patients undergoing drug-eluting stent (DES) implantation for unprotected left main disease (ULMD)., Methods and Results: Consecutive patients who underwent percutaneous coronary intervention for ULMD had prospective platelet reactivity assessment by light transmittance aggregometry after a loading dose of 600 mg of clopidogrel. The primary end point of the study was cardiac mortality, and the secondary end point was stent thrombosis. From January 2005 to September 2008, 215 consecutive patients were treated with DES for ULMD. The incidence of HRPR after clopidogrel loading was 18.6%. The median follow-up was 19.3 months. The overall estimated 1-, 2- and 3-year cardiac mortality rate was 3.9+/-1.3%, 7.5+/-2.2%, and 12.2+/-3.4%, respectively. The 3-year cardiac mortality rate was 8.0+/-3.1% in the low residual platelet reactivity (LRPR) group and 28.3+/-10.4% in the HRPR group (P=0.005). The 3-year stent thrombosis rate was 4.2+/-1.8% in the low residual platelet reactivity group and 16.0+/-7.3% in the HRPR group (P=0.021). By forward stepwise regression analysis, HRPR after clopidogrel loading was the only independent predictor of cardiac death (hazard ratio, 3.82; 95% confidence interval,1.38 to 10.54; P=0.010) and stent thrombosis (hazard ratio, 3.69; 95% confidence interval, 1.12 to 12.09; P=0.031)., Conclusions: HRPR after 600-mg clopidogrel loading is a strong marker of increased risk of cardiac death and DES thrombosis in patients receiving DES stenting for ULMD. Routine assessment of in vitro residual platelet reactivity after clopidogrel loading in patients with ULMD potentially suitable for DES-supported percutaneous coronary intervention should be considered to guide patient care decisions.

Background: The clinical impact of platelet aggregation assessed by point-of-care assays is unknown. We sought to evaluate whether high residual platelet reactivity (RPR) to ADP during clopidogrel therapy, measured by a point-of-care assay, predicts adverse clinical events in acute coronary syndrome patients undergoing percutaneous coronary intervention., Methods and Results: We used the VerifyNow P2Y12 assay (Accumetrics Inc, San Diego, Calif) to determine RPR to ADP in 683 patients with acute coronary syndrome undergoing dual-antiplatelet therapy who underwent percutaneous coronary intervention with bare-metal or drug-eluting stent implantation. All patients received a single 600-mg clopidogrel loading dose followed by 75 mg of clopidogrel daily and 100 to 325 mg of aspirin daily. The end points of the study at follow-up of 12 months were cardiovascular death, nonfatal myocardial infarction (MI), and target-vessel revascularization. At a 12-month follow-up, we found 51 ischemic events (24 cardiovascular deaths [3.5%], 27 nonfatal MIs [3.9%]) and 40 target-vessel revascularizations (5.8%). By receiver operating characteristic curve (ROC) analysis, the optimal cutoff value in predicting 12-month cardiovascular death and nonfatal MI was P2Y12 reaction unit values > or =240. RPR, defined in the presence of P2Y12 reaction unit values above this cutoff, was found to be a significant and independent predictor of cardiovascular death and nonfatal MI in a model that adjusted for cardiovascular risk factors, renal failure, reduced left ventricular ejection fraction, multivessel disease, total stent length, bifurcation lesions, number of lesions treated, type of stent, and use of glycoprotein IIb/IIIa inhibitors (cardiovascular death: hazard ratio 2.55, 95% CI 1.08 to 6.07, P=0.034; nonfatal MI: hazard ratio 3.36, 95% CI 1.49 to 7.58, P=0.004). No significant association was found between high RPR and the risk of target-vessel revascularization., Conclusions: RPR to ADP with clopidogrel therapy, measured by the point-of-care assay VerifyNow P2Y12, is able to detect acute coronary syndrome patients at risk of 12-month cardiovascular death and nonfatal MI. The optimal cutoff value was identified as being 240 P2Y12 reaction units.

Background: Activated clotting time (ACT) is used to monitor heparin therapy during cardiopulmonary bypass, interventional cardiology, and hemodialysis. Traditionally, ACT is performed by use of the Hemochron system. Recently, a new device, the i-STAT system, has been introduced to measure ACT. The aim of this study was to correlate the performances of these two systems and to compare ACT values with heparin levels., Methods: One hundred sixty-five samples from 29 patients undergoing cardiopulmonary bypass or hemodialysis were assayed in duplicate with two Hemochron and two i-STAT devices. Heparin levels were determined by anti-factor Xa assay., Results: The Hemochron ACT ranged from 88 to 1,028 s, and the i-STAT ACT ranged from 80 to 786 s. Heparin plasma levels ranged from 0.01 to 10.8 U/mL. Bland-Altman analysis showed a mean difference between the two methods of 24 +/- 101 s. Strong relationships between anti-factor Xa activity and Hemochron ACTs (r2 = 0.69, P < 0.001) and i-STAT ACTs (r2 = 0.79, P < 0.001) were observed. During cardiac surgery, significant correlations were found: Hemochron, r2 = 0.61, P < 0.001 and i-STAT, r2 = 0.74, P < 0.001. During hemodialysis, relationships between anti-factor Xa activity and ACTs were found: Hemochron, r2 = 0.62, P < 0.001 and i-STAT, r2 = 0.55, P < 0.001., Conclusions: During cardiopulmonary bypass procedure and hemodialysis, i-STAT provides measurements of clotting time quite similar to Hemochron ACT, which were significantly correlated with heparin levels.

Background: We previously demonstrated among renal-transplant recipients (RTRs) a high prevalence of hyperhomocysteinemia, which might account for their elevated cardiovascular risk. The purpose of our study was to document, in hyperhomocysteinemic RTRs, the effect of vitamin supplementation on carotid intima-media thickness (cIMT), which is an early sign of atherosclerosis., Methods: A total of 56 stable hyperhomocysteinemic RTRs were randomly assigned to vitamin supplementation (folic acid 5 mg/day; vitamin B(6) 50 mg/day; vitamin B(12) 400 microg) (group A) or placebo treatment (group B) for 6 months. All subjects underwent cardiovascular risk-factor assessment, including fasting homocysteine (Hcy) levels assay, and high resolution B-mode ultrasound to measure the intima-media thickness of common carotid arteries, at time of enrollment and after 6 months., Results: Fasting Hcy levels markedly decreased in group A after treatment (21.8 [15.5-76.6] micromol/L vs. 9.3 [5.8-13] micromol/L; P<0.0001), whereas no significant changes were observed in group B (20.5 [17-37.6] micromol/L vs. 20.7 [15-34] micromol/L; P=not significant). In group A, cIMT significantly decreased after treatment (0.95+/-0.20 mm vs. 0.64+/-0.17 mm; P<0.0001). All except one patient showed a reduction of cIMT and the mean percentage of cIMT decrease was -32.2+/-12.9%. Patients with methylenetetrahydrofolate reductase (MTHFR) C677T +/+ genotype, with higher Hcy levels, had the major percentage of decrease of Hcy with respect to the other genotypes (mean decrease: MTHFR +/+ 74.8+/-5.7%; MTHFR +/- 58.1+/-10%; MTHFR -/- 56.3+/-8.6%). In hyperhomocysteinemic patients without vitamin supplementation (group B) we documented a significant increase in cIMT after 6 months (0.71+/-0.16 mm vs. 0.87+/-0.19 mm; P<0.05). In 19 of 28 subjects we observed an increase in cIMT, and in 9 of 28 the cIMT was unmodified. The mean percentage of cIMT increase was + 23.3+/-21.1%., Conclusions: Our results demonstrate a beneficial effect of the treatment of hyperhomocysteinemia by vitamin supplementation on cIMT in a group of RTRs.